Abstract

Herpes Simplex Virus (HSV) is a pervasive human pathogen that can establish both symptomatic productive infections and asymptomatic latent infections. During infection, the HSV genome undergoes physical changes that are regulated by cellular and viral proteins. These changes lead to either a template for genome replication during the productive cycle or a persistent stable genome configuration during latency. Changes in viral genomes and events leading to these changes during a particular life cycle are not clearly understood. Using both Gardella gel analysis for circular HSV genomes and restriction enzyme analysis for end-to-end linkages of HSV genomes, we show that HSV genomes circularize only in the absence of the HSV immediate early gene ICP0. In the presence of ICP0 genome circularization is inhibited. Although HSV replication has been previously thought to initiate by a theta mechanism from a circular genome template, these results suggest that HSV replication initiates from a linear genome template due to the presence of ICP0 during lytic infection. We also show that circular genomes are the stable form during long-term persistent infections that model latency. Because HSV genomes begin as linear, double-stranded DNA during infection, host cells may treat the ends of incoming genomes as DNA double strand breaks (DSB) and subsequently repair these ends by circularization of genomes. However, it is unclear if/how these DSB repair pathways contribute to the manipulation of HSV genome configurations during infection and how viral proteins, in particular ICP0, may alter/counteract this repair response to form a template for replication during the productive cycle. Here we show that the cellular DSB repair mechanism, non-homologous end-joining (NHEJ), is the major mechanism by which HSV genomes are circularized. ICP0 not only inhibits HSV genome circularization but also affects the abundance of proteins involved in NHEJ and the distribution of other repair proteins during infection. The study presented here begins to uncover how the interplay between host cell repair responses and the virus' reply to these responses contributes to forming either a genome template for replication during the productive cycle or a persistent stable configuration during latency.